Citrus Extract as a Perspective for the
Control of Dyslipidemia: A Systematic
Review With Meta-Analysis From
Animal Models to Human Studies
Betina M. R. Carvalho 1, Laranda C. Nascimento 1, Jessica C. Nascimento 1,
Vitória S. dos S. Gonçalves 2, Patricia K. Ziegelmann 3, Débora S. Tavares 4 and
Adriana G. Guimarães 5*
1Programa de Pós-Graduação em Ciências Aplicadas à Saúde, Universidade Federal de Sergipe, Lagarto, Brazil, 2Departamento
de Química, Universidade Federal de Sergipe, São Cristóvão, Brazil, 3Departamento de Estatística, Programa de Pós-graduação
em Epidemiologia, Universidade Federal do Rio Grande do Sul, Porto Alegre, Brazil, 4Departamento de Educação em Saúde,
Universidade Federal de Sergipe, Lagarto, Brazil, 5Departamento de Farmácia, Universidade Federal de Sergipe, São Cristóvão,
Brazil
This study aims to obtain scientific evidence on the use of Citrus to control dyslipidemia.
The surveys were carried out in 2020 and updated in March 2021, in the PubMed, Scopus,
LILACS, and SciELO databases, using the following descriptors: Citrus, dyslipidemias,
hypercholesterolemia, hyperlipidemias, lipoproteins, and cholesterol. The risk of bias was
assessed according to the Cochrane methodology for clinical trials and ARRIVE for
preclinical trials. A meta-analysis was performed using the application of R software. A
total of 958 articles were identified and 26 studies demonstrating the effectiveness of the
Citrus genus in controlling dyslipidemia were selected, of which 25 were included in the
meta-analysis. The effects of Citrus products on dyslipidemia appear consistently robust,
acting to reduce total cholesterol, LDL, and triglycerides, in addition to increasing HDL.
These effects are associated with the composition of the extracts, extremely rich in
antioxidant, as flavonoids, and that act on biochemical targets involved in lipogenesis and
beta-oxidation. The risk of bias over all of the included studies was considered critically low
to moderate. The meta-analysis demonstrated results favorable to control dyslipidemia by
Citrus products. On the other hand, high heterogeneity values were identified, weakening
the evidence presented. From this study, one can suggest that Citrus species extracts are
potential candidates for dyslipidemia control, but more studies are needed to increase the
strength of this occurrence.
Keywords: dyslipidemia, citrus, hyperlipidemia, flavonoids, cholesterol
Systematic Review Registration: [https://www.crd.york.ac.uk/prospero/display_record.php?ID=
CRD42019121238], identifier [PROSPERO 2019 CRD42019121238].
INTRODUCTION
Dyslipidemia has high rates of occurrence in the world population (Pirillo et al., 2021), being closely
related to obesity, metabolic syndrome (Mach et al., 2020), atherosclerosis (Wiggins et al., 2019),
Edited by:
Irwin Rose Alencar de Menezes,
Regional University of Cariri, Brazil
Reviewed by:
Amir Hadi,
Isfahan University of Medical
Sciences, Iran
Praveen Kumar M,
Nference, India
*Correspondence:
Adriana G. Guimarães
adrianagibara@hotmail.com
adrianagibara@pq.cnpq.br
Specialty section:
This article was submitted to
Gastrointestinal and Hepatic
Pharmacology,
a section of the journal
Frontiers in Pharmacology
Received: 26 November 2021
Accepted: 10 January 2022
Published: 14 February 2022
Citation:
Carvalho BMR, Nascimento LC,
Nascimento JC, Gonçalves VSS,
Ziegelmann PK, Tavares DS and
Guimarães AG (2022) Citrus Extract as
a Perspective for the Control of
Dyslipidemia: A Systematic Review
With Meta-Analysis From Animal
Models to Human Studies.
Front. Pharmacol. 13:822678.
doi: 10.3389/fphar.2022.822678
Frontiers in Pharmacology | www.frontiersin.org
February 2022 | Volume 13 | Article 822678
1
SYSTEMATIC REVIEW
published: 14 February 2022
doi: 10.3389/fphar.2022.822678
coronary heart disease (Zhao et al., 2021), increased susceptibility
to cancer (Khan et al., 2021), and more recently increased
mortality and severity of COVID-19 (Atmosudigdo et al.,
2021). This disorder is characterized by changes in the lipid
profile, including an increase in total serum cholesterol, low-
density lipoprotein (LDL-c), and triglycerides, as well as a
decrease in high-density lipoprotein (HDL-c) rates in the
blood (Fruchart et al., 2008). The relationships between these
markers have been used as indicators of insulin resistance and
metabolic disorders (Sowndarya et al., 2021), in addition to
atherosclerosis and coronary heart disease (Abid et al., 2021).
However, inflammation markers such as us-CRP (high serum
sensitivity C-reactive protein) can also be considered important
indicators to estimate the severity and risk of coronary artery
disease (Patil et al., 2020). Although there are therapeutic options
for the treatment of dyslipidemias, these are not fully effective,
due to non-adherence to treatment by various factors such as
adverse
effects,
intolerance,
regimen
complexity,
and
imperceptible benefits, besides the need to combine drugs to
improve the clinical condition (Schulz, 2006; Ingersgaard et al.,
2020). On the other hand, lipid-lowering drugs are still
inaccessible to the majority of the population in low-income
countries (Pirillo et al., 2021), making the search for new
strategies to control dyslipidemia necessary.
In this sense, searching for new treatment strategies for this
important health problem is necessary. In this perspective, several
plants and natural products have been studied regarding their
effects on dyslipidemia control (Ballard et al., 2019; Adel
Mehraban et al., 2021); among them, the species of the genus
Citrus (Lamiquiz-Moneo et al., 2020) stand out. Belonging to the
Rutaceae family, the genus Citrus is widely distributed in tropical
and subtropical regions (Manuel et al., 2020) and contains several
substances with biological and nutritional potential, such as fibers
(e.g., pectin), vitamins, and bioactive compounds, with emphasis
on the flavonoids (Alam et al., 2013; Rafiq et al., 2018). Naringin,
naringenin, nobiletin, narirutin, and hesperidin correspond to the
most frequently found flavonoids. They have pronounced
antioxidant and anti-inflammatory activities (Tripoli et al.,
2007; Craft et al., 2012), in addition to being effective in
controlling metabolic syndromes, lipid changes, and obesity
(Geleijnse et al., 1999; Lee et al., 2001; Gattuso et al., 2007;
Alam et al., 2013; Sahebkar, 2017; Ballard et al., 2019).
Thus, this review sought to compile the scientific findings that
demonstrate the effect of Citrus extracts on the control of serum
lipid levels, measuring the size of the effect through meta-analysis.
MATERIAL AND METHODS
Focused Question
The question to be answered was established from the
bibliographic survey “Are species of the genus Citrus effective
in reducing dyslipidemia?” conducted through four steps: (Pirillo
et al., 2021) identification of the use of the Citrus species, (Mach
et al., 2020) identification of the pathology to be applied
(dyslipidemia), (Wiggins et al., 2019) definition of the types of
studies included (preclinical and clinical), and (Zhao et al., 2021)
definition of the target outcome to be analyzed, which is the lipid
profile, building the PICOS strategy (patient or pathology,
intervention, control, other outcomes, and the type of study).
PICOS is highlighted as follows: P: dyslipidemia; I: species of the
genera Citrus (extract); C: untreated or placebo-treated and
hyperlipidemia-induced group; O: blood lipid levels; and S:
preclinical or clinical studies.
Review Writing and Registration of
Protocols
The writing of this systematic review was based on the
recommendations
of
the
Preferred
Reporting
Items
for
Systematic Reviews and Meta-Analyses (PRISMA) (Page et al.,
2021) tool. In addition, the instrument that guides how the
experimental studies should be analyzed was ARRIVE (Animal
Research:
Reporting
of
In
Vivo
Experiments)
guidelines
(Kilkenny et al., 2010). The protocol for this review was
registered
in
the
International
Prospective
Register
Systematic Reviews (Prospero) database and registered on theof
website https://www.crd.york.ac.uk/prospero/, through approved
registry No. CRD42019121238.
Literature Search
The search was carried out through search strategies in the
LILACS, PubMed, SciELO, and Scopus databases in 2019 and
updated in March 2021. The terms used to compose the search in
the databases were defined from consultations with MeSH and
DeCS descriptors. Thus, the following search strategy was
structured: “CITRUS” AND “Lipoproteins” OR “Cholesterol”
OR
“Epicholesterol”
OR
“Dyslipidemias”
OR
“Dyslipoproteinemia” OR “Hypercholesterolemia” OR “High
Cholesterol Levels” OR “Hyperlipidemias” OR “Lipidemia,”
described in detail in Supplementary Table S1.
Study Selection and Eligibility Criteria
After excluding duplicate records, titles, abstracts, and full texts
were independently analyzed by two researchers in order to
determine the study’s eligibility for inclusion in the review.
The inclusion criteria were preclinical studies or randomized
clinical trials that include the use of Citrus species to assess the
effect on the lipid profile. In this review, were excluded reviews,
case studies, case reports, and studies that did not assess the
action on the lipid profile, which included the use of juices from
Citrus species and their action on the lipid profile, or the
association of Citrus species with another compound that
could modify the lipid profile, as well as studies that used
compounds
isolated
from
Citrus
species
to
target
hyperlipidemia. To assess the agreement among researchers,
the statistical test of the Kappa coefficient (K) was applied.
Data Extraction and Risk of Bias
Assessment
Two independent reviewers extracted data from the included
studies. The data from preclinical studies were as follows: Citrus
species, type of extract and part of the plant, composition,
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Carvalho et al.
Citrus Extract as a Perspective for the Control of Dyslipidemia
hyperlipidemia induction model, evaluation methods, treatment,
animal species, and results (all results that were in mg/dL were
converted to mmol/L using the OnlineConversion.com electronic
calculator according to the type of cholesterol). The data from
clinical studies were as follows: Citrus species, type of extract and
part of the plant, composition, study design/location, sample,
criteria for inclusion and exclusion of participants, pathologies,
treatment, and results (all results that were in mmol/L were
converted
to
mg/dL
using
the
OnlineConversion.com
electronic calculator according to the type of cholesterol). All
the outcomes of the experiments carried out in the articles were
extracted for descriptive and inferential analyses.
Through ARRIVE, we apply the following: precise and concise
description of the content of the article in the title, abstract,
explanation of the methodological approach of the introduction,
general and specific objectives, ethical nature of care and use of
animals, study design regarding the number of animals per group,
experimental procedures, information about animals such as sex,
size, weight, and age, housing and breeding, sample size,
statistical
methods,
description
of
results
and
their
interpretation, and study funding.
All clinical studies included in this research were approved for
methodological quality in the risk checklist of Cochrane
randomized for controlled trials (Cochrane Training, 2019).
Items such as generation of random sequence, concealment of
allocation, certification of participants and professionals, as well
as of evaluators, incomplete and selective outcomes, or whether
the study presents any other problem or fraud were used. The
studies considered as having the highest methodological quality
were those related to randomization, blinding, and complete
outcomes.
Meta-Analysis
The studies selected for the meta-analysis had the following
outcomes
analyzed:
total
cholesterol,
LDL,
HDL,
and
triglyceride levels, including the baseline and post-treatment
data from both the control and treatment groups for both
preclinical and clinical studies. In addition to the primary
outcomes,
to
improve
the
understanding
of
the
effects
observed in preclinical studies, the studies were separated into
the following subgroups: route of administration of the extract,
type of animal, type of extract, and parts of the plant used.
For the quantitative analysis of the articles, the studies selected
presented the value of the sample n, mean, deviation, or standard
error for the serum levels of total cholesterol, LDL, HDL, and/or
triglycerides of the treatment and control groups. All data were
tabulated in Excel and later analyzed using the application of R
software. The heterogeneity of the studies was measured using
Cochran’s Q test, using the I2 statistic, which was considered as
heterogeneous when the p value was less than 0.05. The
heterogeneity between the studies was defined using the I2
statistic, which was considered with an unimportant (I2 <
25%), moderate (25% < I2 < 75%), or high degree of
heterogeneity (I2 > 75%) (Higgins and Thompson, 2002). For
heterogeneous studies (I2 > 75%), the following subgroup
analyses were performed: route of administration, type of
animal, parts of the plant used in the extract, type of fruit,
and type of extract. In addition, we performed a sensitivity
analysis,
sequentially
removing
the
individual
studies
to
determine whether any single study affected the overall effect
estimate.
RESULTS
Study Selection and Study Characteristics
During the search process, 958 articles were obtained: 169 from
PubMed, 762 from SciVerse Scopus, 12 from SciELO, and 15
from LILACS. After analyzing the titles, 598 duplicate articles
were removed. After excluding the repeated articles, 360 titles
were screened for analysis according to the inclusion criteria,
from which 329 studies were excluded for not inducing
hyperlipidemia
in
an
animal
model
or
for
not
having
dyslipidemia installed in the case of clinical studies. In
addition, studies with isolated compounds of the Citrus species
or without evaluation of total cholesterol, LDL-C, HDL-C, or
triglycerides were also excluded.
After this design, 31 articles remained, the full texts of which
were analyzed, thus yielding 27 articles that were finally included
in the qualitative synthesis (Figure 1; Tables 1–3). Of these, 22
studies were preclinical trials (Vinson et al., 1998; Bok et al., 1999;
Terpstra et al., 2002; Zulkhairi et al., 2010; Ding et al., 2012; Kang
et al., 2012; Raasmaja et al., 2013; Lu et al., 2013; Kim et al., 2013;
Muhtadi et al., 2015; Dinesh and Hegde, 2016; Shin et al., 2016;
Ashraf et al., 2017; Fayek et al., 2017; Chou et al., 2018; Feksa
et al., 2018; Mir et al., 2019; Sato et al., 2019; Hase-Tamaru et al.,
2019; Ling et al., 2020; Ke et al., 2020; Lee et al., 2020), 3 were
exclusively clinical studies (Gorinstein et al., 2007; Toth et al.,
2015; Cai et al., 2017) and 1 study contained preclinical and
clinical protocols (Mollace et al., 2011) (Figure 1). For the
quantitative synthesis, 25 articles (Vinson et al., 1998; Bok
et al., 1999; Gorinstein et al., 2007; Zulkhairi et al., 2010;
Mollace et al., 2011; Ding et al., 2012; Kang et al., 2012;
Terpstra et al., 2012; Kim et al., 2013; Lu et al., 2013;
Raasmaja et al., 2013; Muhtadi et al., 2015; Dinesh and Hegde,
2016; Shin et al., 2016; Ashraf et al., 2017; Cai et al., 2017; Fayek
et al., 2017; Chou et al., 2018; Feksa et al., 2018; Hase-Tamaru
et al., 2019; Mir et al., 2019; Sato et al., 2019; Ke et al., 2020; Lee
et al., 2020; Ling et al., 2020) were selected. The level of agreement
among the reviewers was 0.470, being considered as moderate.
Tables 2 and 3 show the general characteristics and results of
the preclinical studies, arranged in the chronological order of
publication. Table 4 present the experimental conditions and
results of clinical trials also arranged in the chronological order.
The selected articles were published between 1998 and 2020,
with a predominance of the number of publications in 2013 (n =
3), 2017 (n = 3), 2019 (n = 3), and 2020 (n = 3). These studies were
conducted mainly in China (n = 6; 23.0%) and Korea (n = 5;
19.2%) followed by Italy (n = 2; 7.6%) and Japan (n = 2; 7.6%), in
addition to other countries in which only 1 study was found as
described in Tables 1–3.
In the 26 selected articles, 15 different species of Citrus
were studied in a dyslipidemia model: C. reticulata (n = 4;
15.3%), C. bergamia (n = 3; 11.5%), C. sinensis (n = 3; 13.6%),
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Carvalho et al.
Citrus Extract as a Perspective for the Control of Dyslipidemia
C. junos Tanaka (n = 2; 9.1%), C. grandis (L.) Osbeck also
called C. maxima (n = 3; 11.5%), C. paradise also known as
grapefruit (n = 2; 7.6%), C. unshiu (n = 2; 7.6%), C. sunki Hort.
Ex Tanaka (n = 1; 3.8%), C. aurantium (n = 1; 3.8%), C. mitis
(n = 1; 3.8%), C. limon (n = 1; 3.8%), C. aurantiifolia (n = 1;
3.8%), C. ichangensis (n = 1; 3.8%), Poncirus trifoliata x Citrus
sinensis (n = 1; 3.8%), and C. changshan-huyou (n = 1; 3.8%).
Among the Citrus species used in the preclinical studies, there
was a predominance of six hybrid species in eight studies,
followed by three orange species in eight studies and three
types of lemons in four publications and tangerine species in
four articles. In the clinical studies, on the other hand, there is
a predominance of orange-based bergamot products (C.
bergamia; n = 3 studies) and a study with supplements
containing grapefruit (C. paradise).
\From these species, hydroalcoholic extracts or organic
fractions (n = 20; 86.9%), aqueous extract (n = 1; 4.3%),
and processed fruits (n = 3; 13.0%) were used, which were
incorporated to the diet (n = 14; 60.8%) or administered orally
by gavage (n = 9; 40.9%). In the clinical trials as a whole,
supplementation with encapsulated dry extract was used or
inclusion in the diet. In addition, 21 studies (80.7%) evaluated
the
chemical
composition
of
the
extracts,
with
predominance of compounds belonging to the class ofa
flavonoids,
such
as
naringin,
hesperidin,
neoeriocitrin,
neohesperidin,
nobiletin,
tangeretin,
and
naringenin
(Figure 2).
As
observed
in
Table
1,
the
method
of
inducing
hyperlipidemia in the preclinical studies was by cholesterol-
rich diet or cafeteria-type diet, conducted with rats (n = 12;
52.1%), mice (n = 8; 34.7%), and hamsters (n = 3; 13.0%). Among
the randomized clinical trials (Table 3), the clinical conditions of
the participants were in their entirety dyslipidemia (n = 4; 100%),
associated or not with coronary disease (n = 1, 25%), and
hypertension and glucose intolerance (n = 1; 25%). In the
preclinical and clinical studies, the outcomes evaluated were
the levels of total cholesterol (TC, n = 18; 100%), HDL (n
= 14; 77.7%), LDL (n = 12; 66.7%), VLDL (n = 2; 13.3%), IDL
(n = 1; 5.5%), and triglycerides (TG, n = 17; 94.4%).
From the analysis of the preclinical and clinical studies (Tables
2–4), it was found that the Citrus species were able to significantly
alter the lipid profile in the 26 (100%) studies, decreasing serum
total cholesterol (n = 25; 96.1%), LDL (n = 14; 53.8%),
triglycerides (n = 17; 65.3%), and VLDL (n = 2; 7.6%) and
increasing HDL (n = 4; 15.3%). In the liver, Citrus also
reduced TC and TG (n = 6; 23.0%), lipid accumulation (n =
5; 19.2%), and weight (n = 2; 7.6%). These effects were
accompanied by the maintenance (n = 1; 3.8%) of glutamic-
oxaloacetic transaminase (GOT), glutamic-pyruvic transaminase
(GPT), and alkaline phosphatase (ALP) serum levels or the
FIGURE 1 | Flowchart of the studies included in the qualitative and quantitative synthesis.
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Carvalho et al.
Citrus Extract as a Perspective for the Control of Dyslipidemia
TABLE 1 | Detailed description of the preclinical studies of the effect of Citrus extract on hyperlipidemia included in the systematic review.
References,
country
Extract, plant
part, and
species
Composition
Model
Evaluated
parameters
Treatment protocol
Animal
(n/group)
Vinson et al.,
Hydroalcoholic extract
of whole dried ripe
fruits C. aurantium
25.7% ascorbic acid
Hamster fed on a high-
cholesterol diet
LDL, VLDL
Feed containing 3% of
the extract or 4% of
the extract associated
with ascorbic acid
(57 mmol/kg diet)
daily, for 4 or
10 weeks
Male
1998 (Vinson
et al., 1998)
9.9% flavonoids (quercetin,
hesperidin, naringenin, and
myricetin)
HDL, TC, TG, foam cell
injury
Golden
Syrian
EUA
31.2% protein
in the aorta artery
Hamsters
(n = 10)
3.2% ash
lipid peroxidation
30% carbohydrates
Bok et al.,
Hydroalcoholic extract
of the peel C. reticulata
2.7 g of protein
Rats fed on a high-
cholesterol diet
Plasmatic and hepatic
TC, TG, HDL, LDL
16.7 g/100 g of diet
for 6 weeks
Male
1999 (Bok et al.,
1999)
1.8 g of fat
AIa, fecal neutral sterols,
HMGR, and ACAT
activities in liver tissue
Sprague
Dawley rats
(n = 10)
Korea
1.0 g of ash
20 g of fructose
16.5 g of glucose
8.6 g of sucrose
0.6 g of hesperidin
0.03 g of naringin and 9.67 g
of other sugars
Terpstra et al.,
2002 (Terpstra
et al., 2002)
Peels or waste stream
material of C. limon
-
Hamster fed on a high-
cholesterol diet
BW, FI, and liver weight
Diets containing 3% of
cellulose or lemon
peels or the waste
stream of the lemon
pectin extraction
Male hybrid
Netherlands
TC of plasma and liver
for 8 weeks
F1B Golden
Plasmatic TG, LDL,
HDL, VLDL
Syrian
bile acids, and fecal
sterols
Hamster
(n = 14)
Mollace et al.,
2011 (Mollace
et al., 2011)
Polyphenolic fraction
of C. bergamia Risso &
Poiteau peeled-off
fruits
Neoeriocitrin (77,700 ppm),
naringin (63,011 ppm),
neohesperidin
(72,056 ppm), melitidine
(15,606 ppm), and
brutieridine (33,202 ppm)
High-cholesterol diet-
induced hyperlipemia
BW, TC, LDL, HDL
10 or 20 mg/kg
daily (p.o.)
Male
Italy
TG and glucose
for 30 days
Wistar
Neutral sterols and fecal
bile acids
Rats
(n = 10)
Zulkhairi et al.,
(Zulkhairi et al.,
2010)
Aqueous extract (5%
and 10%) of dried
whole fruits C. mitis
Phenolic compounds
Rats fed on a high-
cholesterol diet
BW, TC, HDL, LDL, TG,
AIb, sdLDLc
5 mg/kg of extract at
5% and 10%
Male
Malaysia
Scavenging activity of
DPPH radicals, reducing
power, lipid
peroxidation (in vitro)
daily (p.o.)
Sprague
Dawley rats
(n = 6)
for 10 weeks
Ding et al.,
Hydroalcoholic extract
of C. ichangensis peel
Naringin, hesperidin,
poncirin, neoeriocitrin
High-fat diet-induced
BWG, FI
Diet supplemented
with 1% of extract, for
8 weeks
Female
2012 (Ding et al.,
2012)
narirutin, neohesperidin,
naringenin, nobiletin,
Obese
TC, TG, LDL, HDL, and
glucose
C57BL/6
mice (n = 7)
China
and tangeretin
Fecal and hepatic TC
and TG; size of EWAT;
mRNA expression of
PPARγ, LXR, and them
target genes in liver
tissue
Kang et al.,
Hydroalcoholic extract
of C
Tangeretin (55.13 mg/g)
High-fat diet-induced
BWG, FI
150 mg/kg/day of
extract (p.o.)
Male
2012 (Kang et al.,
2012)
sunki peel
Nobiletin (38.83 mg/g)
Obese
TC, TG, GPT, GOT, and
LDH, EPAT weight, liver
fat; p-AMPK, p-ACC,
and adiponectin mRNA
expression in EAT.
for 70 days
C57BL/6
mice
(n = 10)
(Continued on following page)
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Citrus Extract as a Perspective for the Control of Dyslipidemia
TABLE 1 | (Continued) Detailed description of the preclinical studies of the effect of Citrus extract on hyperlipidemia included in the systematic review.
References,
country
Extract, plant
part, and
species
Composition
Model
Evaluated
parameters
Treatment protocol
Animal
(n/group)
Korea
Hesperidin (17.11 mg/g)
In mature 3T3-L1
adipocytes: LKB1,
AMPK, ACC, PKA, and
HSL phosphorylation,
CPT-1a gene
expression, and glycerol
release
Rutin (17.02 mg/g)
Sinensetin (4.23 mg/g)
Raasmaja et al.,
2013 (Raasmaja
et al., 2013)
Hydroalcoholic extract
of C. grandis (L.)
Osbeck whole fruits
Naringin at 19%
High-fat diet-induced
BWG, FI
300, 600, or
1,200 mg/kg (p.o.)
daily
Female
Finland
Obese
TG, TC, HDL, glucose,
insulin, ghrelin, GLP-1
for 12 weeks
Zucker
PYY, leptin, and amylin
in plasma
Rats
(n = 10)
Lu et al.,
Hydroalcoholic extract
of Citrange (Poncirus
trifoliata x C. sinensis)
peel or flesh and seed
Bark extract
High-fat diet-induced
obese
BWG, FI, ipGTT, blood
glucose, serum TG, TC,
LDL and HDL, hepatic
TG and TC
Diet supplemented
with 1% w/w of peel
extract
Female
2013 (Lu et al.,
2013)
Neoeriocitrin (14.5 mg/g),
naringin (8.12 mg/g),
neohesperidin (21.1 mg/g),
and poncirin (14.1 mg/g)
Fecal TC and TG,
histological analysis
or 1% w/w of flesh
and seed
C57BL/6
mice (n = 6)
China
Seed extract
of liver tissue
extract, daily
Poncirin (4.85 mg/g)
Neohesperidin (1.87 mg/g)
Naringin (0.87 mg/g)
mRNA levels of PPARγ,
LXR, and their target
genes in liver tissue
for 8 weeks
Kim et al.,
Hydroalcoholic extract
of C. junos Tanaka
peel
Hesperidin (36.3 mg/100 g)
High-fat diet-induced
obese
BWG, FI
Diet supplemented
with 1% and 5% of
extract
Male
2013 (Kim et al.,
2013)
Naringin (11.6 mg/100 g)
TC, TG, glucose, insulin,
leptin, resistin, GOT,
GPT, histological
analysis of liver tissue
for 9 weeks
C57BL/6 J
mice (n = 8)
Korea
Rutin (2.7 mg/100 g)
AMPK phosphorylation
in muscle tissue
Quercetin (1.7 mg/100 g)
and tangeretin (0.7 mg/
100 g)
AMPK and PPARγ
activation in C2C12 and
HEK293 cells,
respectively
Muhtadi et al.,
2015 (Muhtadi
et al., 2015)
Hydroalcoholic extract
of C. sinensis fruit peel
-
High-fat diet-induced
hypercholesterolemia
TC; glucose in rats
125, 250, and
500 mg/kg (p.o.), daily
for 2 weeks
Male
Indonesia
induced by alloxan
monohydrate
After 4-week diet
Wistar rats
(n = 5)
Dinesh and
Hegde, 2016
(Dinesh and
Hegde, 2016)
Hydroalcoholic extract
of C. maxima leaves
Flavonoids, alkaloids,
carbohydrates, glycosides,
saponins, and tannins
Cafeteria diet and
Olanzapine-induced
obesity
BWG, FI
200 and 400 mg/kg
(p.o.), daily for
4 weeks
Female
India
TC, TG, HDL, LDL,
VLDL, GOT, GPT,
glucose
Wistar rats
(n = 6)
Liver weight and TG
Shin et al.,
Hydroalcoholic extract
of C. junos Tanaka
peel
-
Mice fed on a high-
cholesterol diet
BWG, FI
Diet supplemented
with 1% and 5% of the
extract
Male
2016 (Shin et al.,
2016)
TG, TC, HDL, GOT,
GPT, ALP, histological
analysis
for 10 weeks
C57BL/6 J
mice (n = 8)
Korea
of liver tissue
(Continued on following page)
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Carvalho et al.
Citrus Extract as a Perspective for the Control of Dyslipidemia
TABLE 1 | (Continued) Detailed description of the preclinical studies of the effect of Citrus extract on hyperlipidemia included in the systematic review.
References,
country
Extract, plant
part, and
species
Composition
Model
Evaluated
parameters
Treatment protocol
Animal
(n/group)
Expression of PPARα,
FAS, and HMGR in liver
tissue
Lipid accumulation and
expression of p-AMPK,
p-ACC, PPARα, CPT-1,
and HMGR in HepG2
cells
Ashraf et al.,
Hydroalcoholic extract
of C. sinensis peel
-
Rats fed on high-glucose
or cholesterol-rich diet
BWG, FI
Diet supplemented
with 10% Citrus peel
powder (functional)
and 5% peel extract
(nutraceutical), for
8 weeks
Male
2017 (Ashraf
et al., 2017)
TG, TC, LDL, HDL,
glucose, insulin
Sprague
Pakistan
Dawley rats
(n = 6)
Fayek et al.,
Methanolic extract,
hexanic extract,
aqueous homogenate
of C. reticulata
(Mandarin), C. sinensis
(sweet orange), C.
paradise (white
grapefruit), or C.
aurantiifolia (lime) fruit
peels
Nobiletin (%) in hexanic
extracts
Hypercholesterolemia
induced by diet rich in
cholesterol and bile salts
TC
0.1 ml of the
corresponding extract
(p.o.) for 8 weeks
Male
2017 (Fayek
et al., 2017)
Mandarin (10.14%)
TG and glucose
Wistar rats
(n = 6)
Egypt
Sweet orange (3.6%)
White grapefruit (0.9%)
Lime (0.0045%)
Pectin (%) in peel powder
Sweet orange (21.33%)
Lime (19.7%)
While grapefruit (11.66%)
Mandarin (9.14%)
Chou et al., 2018
(Chou et al.,
2018)
Methanolic extract of
C. reticulata
Narirutin (4.52 ± 0.31 mg/g),
hesperidin (9.14 ± 0.32 mg/
g), nobiletin (2.54 ±
0.07 mg/g)
High-fat diet-induced
AST, ALT, triglyceride,
total cholesterol,
glucose, insulin,
HOMA-IR
1% of the
corresponding extract
for 11 weeks
Male
China
Tangeretin (1.67 ±
0.05 mg/g)
obese
C57BL/6 J
mice (n = 8)
Feksa et al., 2018
(Feksa et al.,
2018)
Hydroalcoholic extract
of leaves of C. maxima
Gallic acid, catechin, caffeic
acid, epicatechin, rutin and
isoquercetin, and the major
compounds
High-fat diet and fructose
Blood count, AST, ALT,
triglyceride, total
cholesterol, LDL, HDL,
glucose, urea,
creatinine,
50 mg/kg
Male
Brazil
were caffeic acid (3.71 mg/g)
and catechin (3.65 mg/g
Wistar rats
(n =
Mir et al., 2019
(Mir et al., 2019)
Hydroalcoholic extract
of C. latifolia
-
Hypercholesterolemia
induced by diet rich in
cholesterol
triglyceride, and total
cholesterol
1% of the
corresponding extract
for 4 weeks
Male
Algeria
Wistar rats
(n = 10)
Sato et al., 2019
(Sato et al., 2019)
C. tumida peel powder
Calorie (275 kcal), moisture
(2.9 g), protein (7.4 g), fat
(2.7 g), ash (4.9 g),
carbohydrate (82.1 g), sugar
(28.4 g), fiber (53.7 g),
galacturonic acid (12.2 g),
and sodium (4.3 mg)
High-fat diet
AST, ALT, triglyceride,
total cholesterol, HDL-
C, creatinine, albumin,
calcium, and LDH
C. tumida peel
powder 5% (w/w)
Male
Japan
C57BL/6 J
mice (n = 8)
Tamaru et al.,
2019
(Hase-Tamaru
et al., 2019)
C. unshiu MARC
lyophilized and
powdered
76.1 g carbohydrate, 7.6 g
crude protein, 0.7 g crude
fat, 2.7 g ash, 12.9 g
moisture, 40.9 g
High-fat diet
Total cholesterol,
triglycerides, free fatty
acids, glucose, insulin,
and leptin
2.5%
Sprague
Dawley (SD)
rats (n = 7)
Japan
total fiber, 6.6 g total pectin,
14.4 g hesperidin, and 3.0 g
narirutin
5.0%, or 10.0%
Lee et al., 2020
(Lee et al., 2020)
C. unshiu: dried
extract (CPEW) and
lyophilized (CPEF)
Hesperidin, narirutin, and
synephrine
High-fat diet
AST, ALT, triglyceride,
total cholesterol, and
LDL-C
CPEW: 50 mg/kg;
100 mg/kg
Male
Korea
CPEF: 50 mg/kg;
100 mg/kg
SD rats
(n = 8)
Ling et al., 2020
(Ling et al., 2020)
C. changshan-huyou
Naringin, narirutin, and
neohesperidin
High-fat diet
PTFC: 25 mg/kg;
50 mg/kg; 100 mg/kg
(Continued on following page)
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Carvalho et al.
Citrus Extract as a Perspective for the Control of Dyslipidemia
reduction of GOT, GPT (n = 2; 7.6%), and lactate dehydrogenase
(LDH) (n = 1; 3.8%).
In addition, some Citrus products also reduced body weight
gain (BWG; n = 7; 26.9%), food intake (FI; n = 1; 3.8%), and lipid
accumulation in adipose tissue or cells (n = 3; 11.5%). In human, a
study also demonstrated their effect on the reduction of waist
circumference (WC), waist-to-hip ratio (WHR), and body mass
index (BMI). Taken together, these effects can reduce the risk of
atherosclerosis as shown in three studies (16.6%). However, its
effects on the lipid excretion are still controversial, since two
studies (11.0%) demonstrate increased excretion, two studies
(11.0%) did not identify changes, and only one study (5.5%)
found a reduction in excretion (Table 3). In parallel, some
authors investigated the effect of Citrus-based products on
glucose and their effects on blood glucose reduction (n = 8;
44.4%), insulin increase (n = 2; 11.0%), and glucose uptake in the
cell (n = 1; 5.5%).
In addition, several targets involved in the energy and nutrient
metabolism have been studied. As can be seen in Table 3, some
species
of
Citrus
demonstrated
effects
on
peroxisome
proliferator-activated receptor γ (PPARγ) and peroxisome
proliferator-activated receptor α (PPARα), downmodulating
fatty
acid
synthase
(FAS),
acyl-CoA
oxidase
(ACO),
uncoupling
protein
2
(UCP2),
and
adipocyte
fatty-acid-
binding protein (aP2), besides upregulating CD36 and acetyl-
CoA carboxylase (ACC). They can also act on liver X receptor
(LXR), reducing lipoprotein lipase (LPL), apolipoprotein E
(ApoE),
and
cholesterol
7α-hydroxylase
(CYP7A1)
and
increasing ATP-binding cassette transporter G1 (ABCG1) and
ATP-binding cassette transporter A1 (ABCA1).
The adiponectin signaling pathway also can be involved in the
lipid control. In fact, some Citrus products were able to increase
adiponectin; stimulate the phosphorylation of LKB1, AMP-
activated
protein
kinase
(AMPK),
ACC,
and
carnitine
palmitoyl transferase-1 (CPT-1); and reduce HMGR and
ACAT activities. Their effects on lipolysis were also observed
by the upmodulation of cAMP-dependent protein kinase (PKA)
and hormone-sensitive lipase (HSL), with increase in glycerol.
Besides adiponectin, Citrus seems to act reducing other
adipocytokines, as leptin and resistin, which regulate the
appetite and glucose metabolism and have been associated
with insulin resistance. Their effects were also observed in the
hormones involved with satiety and hunger control, as leptin,
glucagon-like peptide-1 (GLP-1), and ghrelin. Finally, the
antioxidant potential of Citrus has also been demonstrated,
which can offer benefits in reducing lipid oxidation and in the
development of atheromatous plaques.
Methodological Quality/Risk of Bias
The 23 preclinical studies, using the criteria provided by the
ARRIVE guidelines, were analyzed for methodological quality.
The studies showed a percentage of adequacy varying between 50
and 92% (83.82 ± 10.77%), with a greater weakness in the quality
of the methodological description of the studies (Supplementary
Table S2).
As for the clinical studies included in this research and
evaluated by the Cochrane list (Figure 3), all of them had
blinding outcome evaluators and incomplete outcomes. In
addition, 50% of the articles presented low risk of uncertain
bias regarding the criteria of generating a random sequence,
concealment
of
allocation,
blinding
of
the
participants,
reporting of the selective outcome, and other sources of bias
(conflict of interest, based on the source of funding for the study
and method of determination of the sample size).
Meta-Analysis
For the meta-analysis, the preclinical studies measured the level of
total cholesterol [n = 23; 100%; I2 = 99.1% (98.9%; 99.2%)],
triglycerides [n = 20; 87%; I2 = 99.4% (99.3%; 99.5%)], LDL [n =
12; 52.2%; I2 = 99.1% (98.9%; 99.3%)], and HDL [n = 14; 60.9%; I2
= 93.4% (90.6%; 95.4%)]. As for the clinical studies, three clinical
trials with 92, 98, and 237 participants were included in the
TABLE 1 | (Continued) Detailed description of the preclinical studies of the effect of Citrus extract on hyperlipidemia included in the systematic review.
References,
country
Extract, plant
part, and
species
Composition
Model
Evaluated
parameters
Treatment protocol
Animal
(n/group)
AST, ALT, triglyceride,
total cholesterol, LDL-C,
and HDL-C
Golden
hamsters
(n = 12)
China
Ke et al., 2020
(Ke et al., 2020)
C. reticulata Blanco
Nobiletin (98.34 mg/g),
heptamethoxyflavone
(44.26 mg/g), tangeretin
(26.20 mg/g), and
isosinensetin (26.14 mg/g)
High-fat diet
Triglyceride, total
cholesterol, LDL-C, and
HDL-C
0.2 and 0.5% JZE
C57BL/6 J
mice (n = 8)
China
glutamicp.o., intragastric gavage; TC, total cholesterol; TG, triglycerides; LDL, low-density lipoprotein; HDL, high-density lipoprotein; VLDL, very low-density lipoprotein; LDH, lactate
dehydrogenase; GOT, -oxaloacetic transaminase; GPT, glutamic-pyruvic transaminase; EWAT, epididymal white adipose tissue; PPARγ, peroxisome proliferator-activated receptor γ;
FAS, fatty acid synthase; ACO, acyl-CoA oxidase; LXRα, liver X receptor α; LXRβ, liver X receptor β; AMPK, AMP-activated protein kinase; ACC, acetyl-CoA carboxylase; PKA, cAMP-
dependent protein kinase; HSL, hormone-sensitive lipase. GLP-1, glucagon-like peptide-1; PYY, pancreatic peptide YY; BWG, body weight gain; FI, food intake; ipGTT, intraperitoneal
glucose tolerance test; ALP, alkaline phosphatase; FAS, fatty acid synthase receptor; CPT-1, carnitine palmitoyl transferase-1; HMGR, 3-hydroxy-3-methylglutaryl-coenzyme A reductase;
EPAT, epididymal and perirenal adipose tissue; EAT, epididymal adipose tissue.
aThe duration of the experiment is not explicitly informed in the article. AI, atherogenic index.
b[(TC-HDL)/HDL].
c(LDL/HDL); sdLDL, small dense LDL, particle size.
d(TG/HDL).
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Carvalho et al.
Citrus Extract as a Perspective for the Control of Dyslipidemia
TABLE 2 | Outcomes of the preclinical studies included in this systematic review.
Reference
Experimental group (mmol/L)
Control group (mmol/L)
Summary of results
Vinson et al., (Vinson et al.,
1998)
Baseline: TC: 5.84; HDL: 3.31;
TG: 25.1
Baseline: TC: 10.3; HDL: 2.84;
TG: 41.6
↓TC and TG
10 weeks: TC: 6.88; HDL: 1.68;
TG: 27.1
10 weeks: TC: 15.1; HDL: 1.48;
TG: 55.9
↓ lipid peroxidation
↓ atherosclerosis signals (↓ area and density of foam cells),
without changing BW
Bok et al. (Bok et al., 1999)
Baseline
Baseline:
↓ plasma TC
6 weeks: TC: 2.44; HDL: 0.61;
TG: 1.22
6 weeks: TC: 3.8; HDL: 0.57;
TG: 1.12
↓ hepatic TC and TG, without changing HDL, TG, and LDL
plasmatic
↓AI and cholesterol excretion
↓ HMGR and ACAT activities
Terpstra et al. (Terpstra et al.,
2002)
Baseline:
Baseline:
↓ plasma and liver TC, ↓ VLDL + LDL being more effective in
↓VLDL, without changing HDL, ↑ excretion of fecal neutral
sterols and bile acids
8 weeks (lemon peel): TC: 3.51
8 weeks (cellulose): TC: 4.21
without changing BW, FI, and liver weight
8 weeks (waste stream): TC: 3.44
Mollace et al. (Mollace et al.,
2011)
Baseline:
Baseline:
↓TC, LDL, and TG, without changing BW, HDL and glucose
30 days (10 mg): TC: 5.95; LDL: 4.49;
HDL: 0.58; TG: 2.75
30 days: TC: 8.19; LDL: 6.04;
HDL: 0.53; TG: 2.74
↑ fecal neutral sterols and bile acids
30 days (20 mg): TC: 5.00; LDL: 3.90;
HDL: 0.65; TG: 2.74
Zulkhairi et al. (Zulkhairi et al.,
2010)
Baseline (5%)
Baseline: TC: 1.75; LDL: 0.45;
HDL: 0.85; TG: 0.54
↓ TC, LDL, TG
TC: 1.73; LDL: 0.45; HDL: 1.34;
TG: 0.76
4 weeks
↑ HDL
Baseline (10%)
TC: 2.13; LDL: 0.93; HDL: 0.89;
TG: 0.79
↓AI and sdLDL
TC: 1.68; LDL: 0.49; HDL: 1.27;
TG: 0.74
Antioxidant activity, without changing BW
4 weeks (5%)
TC: 1.28; LDL: 0.27; HDL: 1.39;
TG: 0.63
4 weeks (10%)
TC: 1.06; LDL: 0.23; HDL: 1.54;
TG: 0.53
Ding et al. (Ding et al., 2012)
Baseline:
Baseline:
↓ BWG
8 weeks
8 weeks
↓TC and LDL plasmatic
TC: 2.27; LDL: 0.35; HDL: 2.32;
TG: 0.70
TC: 2.65; LDL: 0.46; HDL: 1.95;
TG: 0.70
↓ hepatic TC, TG, glucose, and adipocyte size, without
changing
Plasmatic FI, HDL, and TG and
fecal TC and TG
↓ expression of PPARγ (↓FAS, ACO, and UCP2 and ↑ CD36) ↓
LXR α and β (↓ ApoE, CYP7A1, LPL, and ↑ABCA1)
Kang et al. (Kang et al., 2012)
Baseline:
Baseline:
↓ BWG without changing in FI
70 days
70 days: TC: 4.63; TG: 1.56
↓ TC, TG, LDH, GOT, and GPT
TC: 3.81; TG: 0.94
↓ weight and cell size of EPAT
↓ liver fat
↑ p-AMPK, p-ACC, p-LKB1, and adiponectin
↑ glycerol release
↑ p-PKA and p-HSL
Raasmaja et al. (Raasmaja
et al., 2013)
Baseline (300 mg/kg)
Baseline
Tendency to ↓ TC, glucose, and TG and ↑ HDL
TC: 3.72; HDL: 1.42; TG: 8.34
TC: 3.56; HDL: 1.67; TG: 7.31
↓ GLP-1 and reversing the ↓ of ghrelin, without changing
BWG, FI
Baseline (600 mg/kg)
12 weeks
PYY, leptin, insulin, and amylin
TC: 3.13; HDL: 1.70; TG: 6.27
TC: 4.13; HDL: 0.52; TG: 15.76
Baseline (1,200 mg/kg)
TC: 3.59; HDL: 1.53; TG: 8.11
12 weeks (300 mg/kg)
TC: 4.23; HDL: 0.44; TG: 16.68
12 weeks (600 mg/kg)
TC: 3.62; HDL: 0.80; TG: 12.57
12 weeks (1,200 mg/kg)
TC: 4.36; HDL: 0.80; TG: 17.42
Lu et al. (Lu et al., 2013)
Baseline
Baseline
↓ BWG
(Continued on following page)
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Carvalho et al.
Citrus Extract as a Perspective for the Control of Dyslipidemia
TABLE 2 | (Continued) Outcomes of the preclinical studies included in this systematic review.
Reference
Experimental group (mmol/L)
Control group (mmol/L)
Summary of results
8 weeks (peel)
8 weeks
Improves glucose tolerance and insulin resistance
TC: 2.30; LDL: 0.36; HDL: 2.00;
TG: 0.70
TC: 2.64; LDL: 0.41; HDL: 1.97;
TG: 0.70
↓ serum glucose, TC, and LDL
8 weeks (seed)
↓ hepatic TC and TG, without changing FI, serum HDL, and
fecal TC and TG
TC: 2.43; LDL: 0.41; HDL: 1.87;
TG: 0.74
↓ PPARγ (↓ ap2, FAS); ↓ LXRβ (↓ LPL and ApoE and ↑ ABCG1)
↓ lipid accumulation in liver tissue
Kim et al. (Kim et al., 2013)
Baseline
Baseline:
↓ BWG, glucose, TG, TC, insulin, leptin, and resistin
9 weeks (1%)
9 weeks
↑ glucose uptake
TC: 2.00; TG: 0.85
TC: 2.37; TG: 0.88
↓ liver tissue fat
9 weeks (5%)
↑ PPARγ and AMPK, without changing FI, GOT, and GPT
TC: 1.91; TG: 0.76
Muhtadi et al. (Muhtadi et al.,
2015)
Baseline (125 mg/kg): TC: 4.31
Baseline: TC: 3.77
↓ TC and glucose
Baseline (250 mg/kg): TC: 5.08
2 weeks: TC: 3.27
Baseline 500 (mg/kg): TC: 4.87
2 weeks (125 mg/kg): TC: 1.88
2 weeks (250 mg/kg): TC: 2.13
2 weeks (500 mg/kg): TC: 2.02
Dinesh and Hegde (Dinesh
and Hegde, 2016)
Baseline
Baseline:
↓ BWG and FI
4 weeks (200 mg/kg)
4 weeks
↓ TC, TG, LDL, and VLDL
TC: 79.76; LDL: 54.31; HDL: 40.68;
TG: 104.3
TC: 88.75; LDL: 74.71; HDL:
35.11; TG: 130.0
↑ HDL
4 weeks (400 mg/kg)
↓ GOT and GPT
TC: 75.77; LDL: 51.75; HDL: 43.22;
TG: 98.05
↓ liver weight and TG
↓ glucose
Shin et al. (Shin et al., 2016)
Baseline:
Baseline:
↓ BWG
10 weeks (1%)
10 weeks
↓ TC, LDL, GOT, GPT, ALP, without changing FI, HDL
TC: 2.89; LDL: 1.81; HDL: 0.87
TC: 4.03; LDL: 3.03; HDL: 0.80
↓ liver fat content and weight
10 weeks (5%)
↑ p-AMPK, p-ACC, PPARα, and CPT-1 expression
TC: 2.96; LDL: 1.80; HDL: 0.80
↓ FAS and HMGR expression
↓ lipid accumulation
Ashraf et al. (Ashraf et al.,
2017)
Baseline (powder)
Baseline
Tendency to
TC: 3.34; HDL: 1.19; LDL: 1.67;
TRI: 1.07
TC: 3.30; HDL: 1.17; LDL: 1.63;
TRI: 1.04
↓ BWG and FI
Baseline (extract)
8 weeks
↓ TG, TC, and LDL
TC: 3.32; HDL: 1.21; LDL: 1.62;
TRI: 1.05
TC: 3.81; HDL: 1.17; LDL: 1.85;
TRI: 1.16
↑ HDL
8 weeks (powder)
↓ glucose and ↑ insulin
TC: 3.14; HDL: 1.21; LDL: 1.52;
TRI: 1.01
8 weeks (extract)
TC: 3.03; HDL: 1.24; LDL: 1.44;
TRI: 0.97
Fayek et al. (Fayek et al.,
2017)
Baseline:
Baseline:
Tendency to ↓ TC
Tangerine (alcoholic extract)
Diet
↓ TG and glucose
TC: 2.00; TG: 0.78
TC: 3.92; TG: 2.66
Orange (alcoholic extract)
TC: 3.25; TG: 0.94
Hybrid (alcoholic extract)
TC: 3.95; TG: 0.85
Lime (alcoholic extract)
TC: 5.47; TG: 0.51
Chou et al. (Chou et al., 2018)
Baseline:
Baseline:
Tendency to ↓ TC
11 weeks (1%)
11 weeks (diet)
↓ TG and insulin resistance
TC: 3.85; TG: 0.44
TC: 4.68; TG: 0.85
Feksa et al. (Feksa et al.,
2018)
Baseline
Baseline:
Tendency to
45 days (50 mg/kg)
45 days (diet): TC: 3.34; TG:
3.38; HDL: 0.47; LDL: 1.23
↓ TG, TC, and LDL
TC: 2.12; TG: 2.84; HDL: 0.34;
LDL: 0.61
Mir et al. (Mir et al., 2019)
Baseline
Baseline:
Tendency to
4 weeks (1%)
4 weeks (diet)
↓ TG and TC
(Continued on following page)
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Carvalho et al.
Citrus Extract as a Perspective for the Control of Dyslipidemia
quantitative analyses, which were performed with patients with
dyslipidemia and demonstrated the Citrus effects on the levels of
total cholesterol [I2 = 94.5% (87.3%; 97.6%)], triglycerides [I2
= 95.6% (90.5%; 98.0%)], LDL [I2 = 96.6% (93.0%; 98.4%)], and
HDL [I2 = 81.4% (42.2%; 94.0%)] (in both, n = 3; 100%).
The presentation of the forest graphs was distributed
according to the results of the levels of total cholesterol,
triglycerides, LDL, and HDL for preclinical and clinical
studies. Through the global analysis of preclinical studies, a
reduction of −1.08 mmol/L (95% CI: 1.23; −0.92; Figure 4A)
was found in total cholesterol, equivalent to 41.76 mg/dL; a
reduction of −0.50 mmol/L (95% CI: 0.69; −0.31; Figure 4B)
was found in triglycerides, corresponding to 44.28 mg/dL; and a
reduction of −0.71 mmol/L (95% CI: 0.97; −0.45; Figure 4C) was
found in LDL, what represents 27.45 mg/dL. In addition, an
increase of 0.11 mmol/L in the HDL levels was verified (95%
CI: 0.05; 0.17; Figure 4D), equivalent to 4.25 mg/dL.
As illustrated in Figure 5, in the studies carried out on
humans, the levels (mg/dL) of total cholesterol (MD = −42.03,
95% CI: 73.53; −10.52), triglycerides (MD = −62.41, 95% CI:
110.09; −14.73), and LDL (MD = −37.76, 95% CI: 69.45; −6.06)
were reduced after treating patients with Citrus extracts. In
addition, it was observed that these patients had increased
HDL levels (MD = 5.85, 95% CI: 0.41; 11.28). Although a high
heterogeneity has been observed (I2 > 75%), the synthase of the
results obtained with individual studies favors treatment to the
control of serum lipids. After the analysis of subgroups, high
heterogeneity was still verified and the sensitivity analysis did not
change the result of the general analysis (data not shown).
DISCUSSION
This systematic review compiled data from 25 studies on the
effects of Citrus-based products in the control of dyslipidemia.
Based on the countries where the studies were carried out, most of
them were developed in countries of Asia (such as Korea and
China) and the European Union, in addition to United States and
Egypt, which are among the biggest Citrus product makers in the
world (FAS, 2018). In fact, countries that have greater production
TABLE 2 | (Continued) Outcomes of the preclinical studies included in this systematic review.
Reference
Experimental group (mmol/L)
Control group (mmol/L)
Summary of results
TC: 3.8; TG: 0.9
TC: 5.9; TG: 1.8
Sato et al. (Sato et al., 2019)
Baseline:
Baseline:
Tendency to
4 weeks (5%)
4 weeks (diet)
↓ TG and TC
TC: 3.31; TG: 0.28; HDL: 2.06
TC: 4.39; TG: 0.41; HDL: 2.42
Tamaru et al. (Hase-Tamaru
et al., 2019)
Baseline:
Baseline:
Tendency to
4 weeks (2.5%)
4 weeks (diet)
↓ TG and TC
TC: 2.01; TG: 1.67
TC: 2.27 TG: 2.00
↓ free fatty acids, glucose, insulin, and leptin
4 weeks (5%)
↓ FAS, G6PDH in cytosol, and PAP in microsome
TC: 2.22; TG: 1.63
4 weeks (10%)
TC: 1.72; TG: 2.74
Lee et al. (Lee et al., 2020)
Baseline
Baseline:
Tendency to
8 weeks (CPEW 50 mg/kg): TC: 4.00;
TG: 2.89; LDL: 2.58
8 weeks (diet): TC: 4.00; TG:
2.89; LDL: 2.58
↓ TG and TC
8 weeks (CPEW 100 mg/kg): TC: 3.54;
TG: 2.52; LDL: 2.27
8 weeks (CPEF 50 mg/kg): TC: 4.08;
TG: 2.79; LDL: 2.56
8 weeks (CPEF 100 mg/kg): TC: 3.64;
TG: 2.59; LDL: 2.37
Ling et al. (Ling et al., 2020)
Baseline
Baseline:
Tendency to
4 weeks (25 mg/kg): TC: 32.00; TG:
10.20; HDL: 2.30; LDL: 11.41
4 weeks (diet)
↓ TG, TC, and LDL-C
4 weeks (50 mg/kg): TC: 22.30; TG:
5.30; HDL: 2.83; LDL: 9.83
TC: 41.59; TG: 11.15; HDL:
4.95; LDL: 11.80
4 weeks (100 mg/kg): TC: 21.70; TG:
5.30; HDL: 2.65; LDL: 8.67
Ke et al. (Ke et al., 2020)
Baseline
Baseline:
Tendency to
4 weeks (0.2%): TC: 5.69; TG: 0.28;
HDL: 4.10; LDL: 1.01
4 weeks (diet)
↓ TG, TC, and LDL-C
4 weeks (0.5%): TC: 5.04; TG: 0.28;
HDL: 3.84; LDL: 0.81
TC: 5.62; TG: 0.41; HDL: 4.20;
LDL: 1.20
TC, total cholesterol; TG, triglycerides; LDL, low-density lipoprotein; HDL, high-density lipoprotein; VLDL, very low-density lipoprotein; BW, body weight; HMGR, 3-hydroxy-3-
methylglutaryl-coenzyme A reductase; ACAT, acyl-CoA cholesterol acyltransferase; AI, atherogenic index; FI, food intake; BWG, body weight gain; PPARγ, peroxisome proliferator-
activated receptor γ; FAS, fatty acid synthase; ACO, acyl-CoA oxidase; UCP2, uncoupling protein 2; CD36, cluster of differentiation 36; LXR, liver X receptor; ApoE, apolipoprotein E;
CYP7A1, cholesterol 7α-hydroxylase; LPL, reducing lipoprotein lipase; ABCA1, ATP-binding cassette transporter A1; LDH, lactate dehydrogenase; GPT, glutamic-pyruvic transaminase;
GOT, glutamic-oxaloacetic transaminase; AMPK, AMP-activated protein kinase; ACC, acetyl-CoA carboxylase; PKA; AMP-dependent protein kinase; HSL, hormone-sensitive lipase;
PYY, pancreatic peptide YY; GLP-1, glucagon-like peptide-1; ABCG1, ATP-binding cassette transporter G1; ALP, alkaline phosphatase; CPT-1, carnitine palmitoyl transferase-1; G6PDH,
glucose-6-phosphate dehydrogenase; PAP, phosphatidic acid phosphohydrolase in the microsome.
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Carvalho et al.
Citrus Extract as a Perspective for the Control of Dyslipidemia
of natural resources tend to explore their products more from a
commercial and scientific point of view.
Through the scientific analyses compiled, we can also verify
that species of the genus Citrus have the potential to reduce the
serum levels of total cholesterol (TC), triglycerides (TGs), LDL,
and VLDL and increase HDL. Consequently, Citrus-based
products reduced the body weight, lipid accumulation, and
atherosclerosis risk by the modulation of proteins and genes
involved in the lipid metabolism. Recently, a study with a
standardized extract containing Citrus sinensis L. Osbeck
associated with Citrus limon (Chiechio et al., 2021) also
demonstrated an effect in controlling the levels of total
cholesterol and triglycerides as well as glycemia, possibly due
to its composition rich in anthocyanins, flavonoids, and
hydroxycinnamic acids, reinstating the high potential of Citrus
species in lipid control.
These effects were studied mainly in the animal models of
dyslipidemia induced by cholesterol- or high-fat diets. In these
protocols, lipids ingested are initially degraded by intestinal lipase
and, in enterocytes, TGs are resynthesized and associated with
cholesterol and lipoproteins (ApoB-48, ApoE, and ApoC-II),
forming chylomicrons. These distributed fatty acids between
tissues and their remnants are metabolized in the liver. In this
organ, fatty acid and glucose activate metabolic pathways for
energy synthesis and storage, so that excess citrate is converted by
citrate lyase (ACLY) into acetyl-CoA, which by the action of
acetyl-CoA
carboxylase
(ACC)
forms
malonyl-CoA.
This
metabolic intermediate is used by the cell to produce fatty acid
through the action of the enzymes Stearoyl-CoA Desaturase-1
(SCD1)
and
fatty
acid
synthase
(FAS),
in
addition
to
downregulating CPT-1, an important transporter of Acil-Coa
into the mitochondria which enables its β-oxidation. These fatty
acids give rise to triglyceride molecules. In addition, acetyl-CoA
can participate in the synthetic pathway of cholesterol, forming
HMG-CoA which is converted into mevalonic acid by HMGR.
This originates the free cholesterol molecule, which can be
TABLE 3 | Detailed description of the clinical studies of the effect of Citrus extract on hyperlipidemia included in the systematic review.
References/
country
Extract, plant
part and
species
Composition
Sample
Pathology
Parameters
evaluated
Treatment protocol
Gorinstein et al.,
Fresh fruit peels of
red grapefruit or
blond grapefruit
processed
Anthocyanins
57 patients
(39–72 years)
Hypertriglyceridemia and
coronary
HR, BP, BW
Daily supplementation with
red or blond grapefruits
associated with anti-
atherosclerosis diet for
30 days (n = 19/group)
2007 (Gorinstein
et al., 2007)
Red: 51.5 mg/100 g
disease
CT, LDL, HDL,
Israel
Blond: 49.3 mg/100 g
TG, serum
antioxidant activity
by ABTS and
TEAC
Flavonoids (naringin)
Red: 21.61 mg/100 g
Blond: 19.53 mg/100 g
Total fibers
Red: 1.39 g/100 g
Blond: 1.37 g/100 g
Mollace et al.,
2011 (Mollace
et al., 2011)
Polyphenolic
fraction of C.
bergamia peeled-
off fruits
Neoeriocitrin (77,700 ppm)
237 patients
Hyperlipemia associated or
not with hyperglycaemia
TC, LDL, HDL,
500 or 1,000 mg/day
encapsulated with 50 mg
ascorbic acid, for 30 days
(n = 104–32/group)
Italy
Naringin (63,011 ppm)
TG, reactive
vasodilation
Neohesperidin
(72,056 ppm) and
melitidine (15,606 ppm)
Brutieridine (33,202 ppm)
Toth et al., 2016
(Toth et al.,
2015)
Bergavit
®
(Bergamot juice
derived extract, C.
bergamia)
150 mg of flavonoids
80 individuals
(42 men and
38 women)
Moderate
hypercholesterolemia
TC, LDL, HDL, TG,
VLDL, IDL, IMT,
LDL size
150 mg/day for 6 months
(n = 80)
Italy
16% of neoeriocitrin
47% neohesperidin
37% naringin
Cai et al., 2017
(Cai et al., 2017)
C. bergamia extract
(CitriCholess
®)
25% bioflavonoids, sterols
and orange oil (820 mg/
day), vitamin C (50 mg/
day), vitamin B6 (20 mg/
daily), B12 (2,000 µg/day),
and folic acid (800 µg/day)
98 older
people
Dyslipidemia and arterial
hypertension and problems
of glucose intolerance
TG, TC, LDL, HDL,
glucose, BW, WC,
HC, WHR,
and BMI
500 mg/day for 12 weeks
(n = 48–50/group)
China
Legend: TC, total cholesterol; TG, triglycerides; LDL, low-density lipoprotein; HDL, high-density lipoprotein; TEAC, Trolox-equivalent antioxidant capacity; HR, heart rate; BP, blood
pressure; BW, body weight; IMT, carotid intima-media thickness; BW, body weight (kg); WC, waist circumference (cm); HC, hip circumference (cm); WHR, waist-to-hip ratio; BMI, body
mass index.
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Carvalho et al.
Citrus Extract as a Perspective for the Control of Dyslipidemia
esterified by acyl-CoA:cholesterol acyltransferase (ACAT) or
converted into bile acids by CYP7A1. TG, free cholesterol, and
cholesterol ester conjugate with lipoproteins (ApoE, ApoC-II,
and ApoB-100) constituting the VLDL molecule (TGs >
cholesterol). This lipoprotein distributes fatty acids to tissues
by the action of lipoprotein lipase (LPL) and becomes IDL (TGs ≈
cholesterol, ApoB-100, ApoE) and later LDL (TGs, < cholesterol,
ApoB-100). That way, high-lipid diets increase the plasmatic
concentrations of TG, TC, VLDL, IDL, and LDL (DiNicolantonio
and O’Keefe, 2018; Andreadou et al., 2020). These mechanisms
can be observed in Figure 6 (black lines).
Through this review, it was found that the effect of Citrus-
based products on the release of adipocytokines and their
signaling pathways has been studied. These molecules are
produced by adipose tissue and control several metabolic
pathways, in addition to affecting the state of hunger and
TABLE 4 | Outcomes of the clinical studies included in this systematic review.
Reference
Experimental group
(mg/dL)
Control group (mg/dL)
Summary of results
Gorinstein et al. (Gorinstein et al.,
2007)
Baseline:
Baseline:
Red: ↓ TC, LDL, and TG
Red
TC: 306.26
Blond: ↓ LDL only
TC: 258.70
LDL: 243.23
Both: ↑ serum antioxidant activity, without change in HR,
BP, BW,
LDL: 193.73
HDL: 46.20
HDL
HDL: 52.59
TG: 205.49
TG: 149.68
Blond
TC: 283.06
LDL: 217.32
HDL: 50.27
TG: 193.97
Mollace et al. (Mollace et al., 2011)
Baseline (500 mg)
Treated with capsules containing
↓ TC, TG, and LDL
TC: 286.00
500 mg of maltodextrin and 50 mg of
ascorbic acid
↑ HDL
LDL: 184.96
Baseline
↓ glucose
HDL: 34.55
TC: 275.67
↑ reactive vasodilation
TG: 266.87
LDL: 186.31
Baseline (1,000 mg)
HDL: 34.59
TC: 279.40
TG: 275.62
LDL: 189.70
TC: 279.40
HDL: 32.78
LDL: 185.64
TG: 270.11
HDL: 35.05
After 30 days (500 mg)
TG: 275.71
TC: 211.42
LDL: 132.79
HDL: 40.53
TG: 180.18
After 30 days (1,000 mg)
TC: 201.99
LDL: 125.34
HDL: 46.00
TG: 157.48
Toth et al. (Toth et al., 2015)
Baseline
Baseline:
↓ TC, LDL, TG, and IMT
TC: 224.28
TC: 255.22
↑ HDL, IDL, and LDL size
LDL: 143.07
LDL: 177.88
without changing VLDL
HDL: 54.13
HDL: 50.27
TG: 132.86
TG: 159.43
Cai et al. (Cai et al., 2017)
Baseline
Baseline
↓ LDL
TC: 211.13
TC: 217.32; LDL: 138.43; HDL: 51.81; TG:
170.94
↓ BW, WC,
LDL: 131.09
TC: 210.36
WHR, and BMI
HDL: 49.88
LDL: 132.63
without changing TG, TC, HDL, glucose, HC
TG: 192.20
HDL: 52.20; TG: 172.71
500 mg
TC: 198.76
LDL: 121.03
HDL: 50.27
TG: 162.09
Legend: TC, total cholesterol; TG, triglycerides; LDL, low-density lipoprotein; HDL, high-density lipoprotein; TEAC, Trolox-equivalent antioxidant capacity; HR, heart rate; BP, blood
pressure; BW, body weight; IMT, carotid intima-media thickness; BW, body weight (kg); WC, waist circumference (cm); HC, hip circumference (cm); WHR, waist-to-hip ratio; BMI, body
mass index.
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Carvalho et al.
Citrus Extract as a Perspective for the Control of Dyslipidemia
satiety and being related to the development of coronary diseases
and metabolic disorders (Cao, 2014). Citrus products reduce
adiponectin (Kang et al., 2012), whose action on specific
receptors (AdipoR) increases the phosphorylation of LKB1 and
AMPK (Kang et al., 2012; Shin et al., 2016). It negatively
modulates ACC (Kang et al., 2012; Shin et al., 2016), reducing
malonyl-Coa levels and, consequently, increasing CPT-1 (Shin
et al., 2016); in addition, it decreases the HMGR activity (Bok
et al., 1999; Shin et al., 2016) and modulates genes like LXR (Ding
et al., 2012; Lu et al., 2013) and PPAR (Kim et al., 2013; Shin et al.,
2016; Lu et al., 2018). Through these genes, Citrus regulates
several protein targets involved in lipogenesis (FAS, aP2, ACC)
(Ding et al., 2012; Lu et al., 2013; Shin et al., 2016), lipoprotein
formation and metabolism (ApoE, LPL) (Ding et al., 2012; Lu
et al., 2013), cholesterol metabolism (CYP7A1) (Ding et al.,
2012), and cholesterol and lipid efflux (ABCG1 and ABCA1)
(Ding et al., 2012; Lu et al., 2013). At the same time, its ability to
stimulate the PKA-HSL pathway has also been observed (Kang
et al., 2012), increasing the degradation of TG in glycerol and fatty
acid, in addition to reducing the activity of ACAT (Bok et al.,
1999), which contributes to the reduction of cholesterol ester
levels. It is worth mentioning that bio-products based on Citrus
help in glycemic control (Mollace et al., 2011; Ding et al., 2012;
Kim et al., 2013; Lu et al., 2013; Raasmaja et al., 2013; Muhtadi
et al., 2015; Dinesh and Hegde, 2016; Ashraf et al., 2017; Fayek
et al., 2017), possibly by reducing resistin (Kim et al., 2013), an
adipocytokine whose increase has been associated with insulin
resistance, atherosclerosis, oxidative stress, and inflammation. All
of these molecular events result in decreased lipogenesis and
increased lipid oxidation, contributing to the control of the lipid
profile (Figure 6).
However, some results seem contradictory, such as the effect
of Citrus in reduction of the mRNA levels of PPARγ target genes,
including ACO and UCP2 in the liver tissue (Ding et al., 2012).
FIGURE 2 | Chemical structure of the main flavonoids found in Citrus.
FIGURE 3 | Methodological quality of clinical trials included.
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Carvalho et al.
Citrus Extract as a Perspective for the Control of Dyslipidemia
ACO is the first enzyme of peroxisomal β-oxidation which will
reduce the accumulation of lipids in the liver and promote its
excretion (Ferdinandusse et al., 2007). On the other hand, UCP2
is an uncoupling protein which acts as a carrier of protons present
in the inner membrane of mitochondria and contributes to
thermogenesis, being a positive factor for the prevention of
obesity (Brand and Esteves, 2005). Thus, upregulation of these
mRNAs would contribute to the observed outcomes. However,
the absence of baseline conditions for these targets makes it
difficult to understand these data, so further studies are needed to
elucidate this mechanism.
Similarly, Citrus seems to increase CD36 (Ding et al., 2012),
the fatty acid translocase protein that facilitates the transport of
fatty acids, the hepatic uptake of fatty acids, and the
accumulation of fat and has a high affinity for binding with
the oxidized LDL molecule, increasing the inflammatory activity
and
being
a
main
condition
for
the
development
of
atherosclerosis
and
thrombosis
(Pepino
et
al.,
2014).
However, the correlation with the observed outcomes also
needs to be further investigated, since the experimental
conditions of the study do not allow a thorough analysis of
this target in the experimental model used, as well as in the
primary outcome studied.
It is also worth noting that some studies have shown that
Citrus can help control hunger promoting the modulation of
ghrelin. Known as “Hunger Hormone,” this peptide is
produced by endocrine cells present in the stomach and
acts in the control of hunger, adiposity, and glucose- and
energy-homeostasis, among other functions (Pradhan et al.,
2013). More over, Citrus also downregulates leptin and GLP-1
levels, which are involved with satiety control. Leptin, a
hormone produced by adipose tissue, plays an important
role in the control of energy homeostasis, the excess and
resistance of which are associated with obesity, leading to
failures
in
the
signaling
mechanisms
associated
with
decreased nutrition and body weight control (Pan and
Myers, 2018). On the other hand, glucagon-like peptide 1
(GLP-1) is a gut hormone that promotes satiety; potentiates
insulin release and suppression of glucagon release in response
to nutrient intake; and decreases postprandial plasma levels of
glucose (Andersen et al., 2018). Thus, the effects observed for
Citrus in the reduction of GLP-1 may be related to overnight
fasting
or
long-term
regulation
of
eating
and
energy
metabolism, requiring further investigation.
The notations are as follows: ABCA1: ATP-binding cassette
transporter A1; ABCG1: ATP-binding cassette transporter G1;
ACAT: acyl-CoA:cholesterol acyltransferase; ACC: acetyl-
CoA
carboxylase;
ACLY:
citrate
lyase;
ACO:
acyl-CoA
oxidase;
AdipoR:
adiponectin
receptor;
AMPK:
AMP-
activated protein kinase; aP2: adipocyte fatty-acid-binding
protein;
ApoB-100:
apolipoprotein
B-100;
ApoC-II:
apolipoprotein C2; ApoE: apolipoprotein E; CD36: cluster
of differentiation 36; CPT-1: carnitine palmitoyl transferase-
1; CYP7A1: cholesterol 7α-hydroxylase; FAS: fatty acid
synthase;
GLUT
4:
glucose
transporter
4;
HMGR:
3-
hydroxy-3-methylglutaryl-coenzyme
A
reductase;
HSL:
FIGURE 4 | Forest plot of the preclinical studies that evaluated the effect
of Citrus species on total cholesterol (A), triglycerides (B), LDL (C), and HDL
(D) levels. The numbers on the x-axis indicate the effect of the treatment and
its favoring. SD: standard deviation of the differences. MD: difference
between the means.
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Carvalho et al.
Citrus Extract as a Perspective for the Control of Dyslipidemia
hormone-sensitive
lipase;
IDL:
intermediate
low-density
lipoprotein;
LDL:
low-density
lipoprotein;
LKB1:
liver
kinase B1; LPL: lipoprotein lipase; LXR: liver X receptor;
p-ACC:
phosphorylated
acetyl-CoA
carboxylase;
PKA:
cAMP-dependent
protein
kinase;
PPAR:
peroxisome
proliferator-activated
receptor;
SCD1:
Stearoyl-CoA
FIGURE 5 | Forest plot of the clinical studies that evaluated the effect of Citrus species on total cholesterol (A), triglyceride (B), LDL (C), and HDL (D) levels. The
numbers on the x-axis indicate the effect of the treatment and its favoring. SD: standard deviation of the differences. MD: difference between the means.
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Carvalho et al.
Citrus Extract as a Perspective for the Control of Dyslipidemia
Desaturase-1; TC: total cholesterol; TGs: triglycerides; UCP2:
uncoupling protein 2; VLDL: very low-density lipoprotein.
The effects of Citrus bioproducts on the lipid profile may be
related to the presence of bioactive compounds, with emphasis on the
flavonoids, such as naringin, hesperidin, neohesperidin, neoeriocitrin,
nobiletin, tangeretin, and naringenin as compiled in this review. In
fact, these compounds are believed to play a very significant role in
reducing the levels of total cholesterol, triglycerides, and LDL
(Mulvihill and Huff, 2012; Assini et al., 2013; Kou et al., 2017;
Zeka et al., 2017). Several studies have shown that naringin reduces
the HMGR activity more potently than does vitamin E (Choi et al.,
2001; Lee et al., 2001), as well as decreasing the action of ACAT (Kim
et al., 2006), which contributed to hypocholesterolemic action and
higher excretion of fecal sterols (Jeon et al., 2004). Similarly,
hesperidin reduces plasma cholesterol in hypercholesterolemic rats
by decreasing ACAT and HMGR (Lee et al., 1999; Lee et al., 2012)
besides changing the expressions of genes encoding PPARs and the
LDL receptor (Akiyama et al., 2009). A recent study demonstrated
that neohesperidin is also able to regulate the lipid metabolism in vivo
and in vitro via FGF21 and AMPK/SIRT1/PGC-1α signaling axis
(Wu et al., 2017). Furthermore, the non-glycoside Citrus flavonoid,
naringenin, stimulates the hepatic fatty acid oxidation via PPARγ and
prevents lipogenesis in both the liver and the muscle, reducing the
serum lipid levels (Mulvihill et al., 2009).
FIGURE 6 | Biochemical and tissue changes caused by diets high in fat and calories (black lines) and mechanisms of action of Citrus products upon metabolic
disorders associated with hyperlipidemia (blue lines indicate activation and red lines indicate inhibition).
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Carvalho et al.
Citrus Extract as a Perspective for the Control of Dyslipidemia
In this review, we also observed that the Citrus products act by
reducing the atherogenic index or tissue manifestations associated
with atherosclerosis (Vinson et al., 1998; Bok et al., 1999; Zulkhairi
et al., 2010). In fact, the polyphenolic compounds and flavonoids
found in the Citrus species have antioxidant (Vinson et al., 1998;
Gorinstein et al., 2007; Zulkhairi et al., 2010; Craft et al., 2012) and
anti-inflammatory properties, in addition to their ability to decrease
LDL levels, inhibiting the formation of atherosclerotic plaques (Tripoli
et al., 2007; Assini et al., 2013; Onakpoya et al., 2017). Naringin, for
example, reduces plaque progression once it decreases non-high-
density lipoprotein cholesterol concentrations and biomarkers of
endothelial dysfunction and inhibits the expression of ICAM-1 in
endothelial cells, preventing immune cell adhesion and infiltration in
the vascular wall (Choe et al., 2001; Chanet et al., 2012).
Confirming the results of the systematic review, the meta-analysis
of preclinical studies indicated that Citrus products reduce the total
cholesterol, triglycerides, and LDL levels by −41.76, −44.28, and
−27.45 mg/dL, respectively, while increasing the HDL levels by
4.25 mg/dL. Similar results were observed in the clinical studies, in
which the Citrus species induce a reduction in the total cholesterol,
triglycerides, and LDL levels by −42.03, −62.41, and −37.76 mg/dL,
respectively, whereas the HDL levels increased by an average of
5.85 mg/dL.
In the meta-analysis published by Onakpoyaa et al. (2015)
(Onakpoya et al., 2017), performed with two clinical trials
about the effect of grapefruits on the lipid profile, significant
effects were observed only for the increase in HDL, without TC
and LDL changes. More recently, a meta-analysis published by
Kou et al. (2017) showed that the sizes of effect measures for
LDL and total cholesterol presented significant results in the
group
of
patients
treated
with
Citrus
juice,
without
considerable changes in HDL and TG levels. The divergence
between the results presented in our meta-analysis compared
to those previously published is justified by the broader scope
of our question, as well as the inclusion of more recent studies,
which have confirmed the contribution of Citrus-based
products in the control of blood lipids.
Through the analysis of the risk of bias, it can be observed
that the preclinical studies have a satisfactory average score,
with some limitations in the methodological description of the
studies
and
the
results.
Similarly,
clinical
studies
had
limitations
in
reporting
or
methodology
in
terms
of
blinding, allocation, randomization, and reporting of results.
The use of tools to assess the risk of bias in the studies included
in the systematic reviews has been widely well supported by
groups such as SYRCLE (Hooijmans et al., 2014), ARRIVE
(Kilkenny et al., 2010), and Cochrane (Cochrane Training,
2019), since the credibility of the results and the strength of the
evidence depend on the methodological criteria of the studies
(Busch et al., 2020).
Thus, although the results obtained are favorable to the treatment
with Citrus extracts, the methodological limitations and high
heterogeneity of the studies included in the meta-analysis weaken
the evidence about the real benefits of this intervention. In addition,
the studies do not provide information on effective dose,
bioavailability, efficacy, and safety. These parameters are required
to propel the use of these promising therapeutic agents into the
clinical area. For this reason, further studies are needed to strengthen
the evidence of the effects of Citrus on dyslipidemia.
This systematic review presents as limitations the low evidence
found due to the high variability of the studies and variation of the
methodological protocols of the articles. Among them, we can
mention the differences in the induction of dyslipidemia, routes
of administration, and types of extracts, besides the absence of
baseline serum levels of lipids for comparison after the induction
and inconclusive report. Finally, as in our review, of the 25 studies
included in the meta-analysis, only 3 presented results in humans; we
chose not to use the GRADE system. For this reason, we believe that
further clinical studies are needed to provide sufficient scientific
support
to measure
the
effectiveness of
Citrus
effects
on
dyslipidemia.
CONCLUSION
From the compilations of the studies, one can suggest that the
Citrus extract has a potential effect in dyslipidemia control, both
in the preclinical studies and clinical trials. These effects can be
associated with the presence of bioactive compounds, as
flavonoids, which act synergistically through several pathways,
causing inhibition of lipogenesis and activating β-oxidation.
However, due to the high heterogeneity of the reposted
findings, further studies are needed to increase the strength of
clinical evidence of the action of Citrus extracts on the control of
dyslipidemia and increase the strength of that evidence.
DATA AVAILABILITY STATEMENT
The original contributions presented in the study are included in
the article/Supplementary Material; further inquiries can be
directed to the corresponding author.
AUTHOR CONTRIBUTIONS
Ideation and preparation of the review: BC and AG; search and
selection of studies: BC and LN; third evaluation for discrepancy
analysis: AG; qualitative data extraction: BC, LN, and JN; quantitative
data extraction: BC and VG; meta-analysis: BC, VG, and PZ; writing
and finalizing the review: BC, DT, and AG.
FUNDING
The authors acknowledge grants from the Foundation for Support
of Research and Technological Innovation of the State of Sergipe
(Fundação de Apoio à Pesquisa e Inovação Tecnológica do Estado de
Sergipe: FAPITEC/SE, EDITAL CAPES/FAPITEC/SE N° 10/2016 –
PROMOB
1995/2017),
National
Council
for
Scientific
and
Technological
Development
(Conselho
Nacional
de
Desenvolvimento
Científico
e
Tecnológico:
CNPq/Brazil),
Coordination
for
the
Improvement
of
Personnel
Higher
Education (Coordenação de Aperfeiçoamento de Pessoal de Nível
Superior: CAPES/Brazil), and Federal University of Sergipe.
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February 2022 | Volume 13 | Article 822678
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Carvalho et al.
Citrus Extract as a Perspective for the Control of Dyslipidemia
ACKNOWLEDGMENTS
We are grateful for the support given by Patrícia K. Ziegelmann in
the elaboration of the meta-analysis and to teacher Abilio Borghi
for the assistance with English language review.
SUPPLEMENTARY MATERIAL
The Supplementary Material for this article can be found online at:
https://www.frontiersin.org/articles/10.3389/fphar.2022.822678/
full#supplementary-material
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Conflict of Interest: The authors declare that the research was conducted in the
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Carvalho et al.
Citrus Extract as a Perspective for the Control of Dyslipidemia